Title: Cholinergic brainstem signaling in striatal circuits, PI: Juan Mena-Segovia PROJECT SUMMARY The striatum is the main input structure of the basal ganglia and it plays a central role in action reinforcement, movement planning and execution of motor sequences. Two neuromodulators exert powerful control over striatal neuronal circuits: dopamine and acetylcholine. These neuromodulatory systems are inextricably linked, such that they are anatomically interconnected and reciprocally regulated. A significant number of neurological disorders that affect the basal ganglia are associated with the dysregulation of one or both of these neuromodulatory systems (e.g. Parkinson?s disease, Tourette syndrome). Understanding how these systems are regulated and how they modulate each other is fundamental for understanding normal basal ganglia function and how they are altered in the diseased brain. Until recently, striatal acetylcholine was believed to originate exclusively from the striatal cholinergic interneurons. However, we discovered an extrinsic source of acetylcholine to the striatum originating in the pedunculopontine nucleus (PPN) and laterodorsal tegmental nucleus (LDT) of the brainstem. Dopamine, on the other hand, originates from the midbrain and innervate large extents of the striatum, and the activity of dopamine neurons is strongly modulated by the axon collaterals of cholinergic PPN/LDT neurons. Our preliminary data using optogenetic tools show that PPN and LDT produce a robust and direct modulatory effect on distinct types of striatal neurons, including cholinergic interneurons. Thus, PPN/LDT neurons are not only able to modulate the activity of dopamine neurons that in turn project densely to the striatum, but they are also capable of directly modulating striatal microcircuits. Our data thus suggest that the PPN and LDT are in a key position to modulate striatal function. The goal of the proposed studies is to characterize the anatomical organization and functional significance of the brainstem cholinergic innervation of the striatum. First, we will identify the striatal domains that are preferentially innervated by brainstem cholinergic axons and identify the subtypes of striatal neurons targeted by PPN and LDT axons. Second, using optogenetic methods, we will characterize the impact of PPN and LDT axons on neurochemically identified single striatal neurons. Third, we will identify the direct impact of the PPN and LDT cholinergic neurons on striatal-dependent behaviors. Furthermore, because striatal cholinergic interneurons and PPN/LDT cholinergic neurons are two markedly distinct populations in terms of their synaptic inputs, intrinsic physiological properties and pattern of activation during behavior, we expect to see critical differences in how they influence striatal circuits and striatal function. Thus, we will compare the anatomical and physiological organization of these two sources of acetylcholine. The discovery of an extrinsic source of acetylcholine to the striatum highlighted a major gap in our understanding of how neuromodulators regulate striatal activity. Current basal ganglia models are incomplete and need to be revised to incorporate a new major player in basal ganglia function: the cholinergic brainstem. Addressing these issues is not only timely and of critical importance for the progress of basal ganglia research, but for understanding striatal-dependent neuropathology.